Abstract
Human fibroblasts were exposed to sodium arsenite for 4h. The cytotoxicity of sodium arsenite treatment was determinted by SRB assay. The IC50 was 30μM- 40μM. Here, the cell survival was only 20﹪of control in the exposure of 80μM sodium arsnite. This study has shown that sodium arsnite influence the cytotoxicity of HF. Treatment of HF with SA40-160μM for 4h increase the nitrite content released into cell cultured medium,indicating the elevated level of nitric oxide (NO). Peroxynitrite is a powerful modifier of intracellular proteins. The nitration of tyrosine, nitrotyrosine, is a useful marker for detecting peroxynitrite in biological samples.HF cells exposure to SA40-160μM for 4h increase nitrotyrosine level by 10-20﹪,indicating the elevated level of peroxynitrite. Cotreament with 3.2mM Nω-nitro-L-arginine methyl ester ( NAME) and 10μM SA for 4h, the survival rose from 65﹪to 78﹪. These data suggest that NO was involed in SA-induced cell death. HF cells incubated with 40-160μM SA showed glutathione reductase activity significantly decrease to less than 29-60﹪after 2h and less than 43-65﹪after 4h. After HF cells were exposed to 40μM SA for 4h and followed a postincubation in drug-free medium for 12 or 24h. Glutathione reductase(GR) activity rose from 20﹪to 30﹪after 12 and 24 h of recovery. However,GR activity did not recover to the control level wthin 24h. Cotreament of HF cells with SA and NAME（1.6 or 3.2mM） for 4h, partly restore the GR activity. At 1.6mM NAME,the percentage of the recovered GR activity was 4﹪-10﹪.At 3.2mM NAME,the percentage of the recovered GR activity was 4﹪-30﹪. The data showed that treatment with SA increased nitrite production in HF cells and that NAME could not significantly recover GR activity. HF cells exposure to SA40-160μM for 4h decrease GSH level by 16-56nmol /mg protein. This decrease in GSH content was accompanied by a simultaneous increase in GSSG level, with a GSH disappearance and GSSG production.4h after an incubation with 40μM SA, GSSG increase about 2.6-fold.Intracellular total glutathione concentration were almost unchanged suggesting that GSH synthesis did not differ from control cells after 4h. Cotreament with GR and 10μM SA for 4h, the survival rose from 65﹪to 82﹪. These data suggest that decreased glutathione reductase induced by sodium arsenite, lead to cell loss of viability.